Differential activation of extracellular signal-regulated kinase 1 and a related complex in neuronal nuclei

Abstract

The Extracellular signal-Regulated Kinases 1 and 2 (ERKs 1/2) are known to participate in regulating transcription in response to moderate depolarization (synaptic stimulation), but how the same active enzyme can differentially regulate distinct transcriptional programs induced with abnormal depolarization (high potassium) is unknown. We hypothesized that ERK1 or 2 accomplishes this differential nuclear response through close association with other proteins in stable complexes. In support of this hypothesis, we have found that immunoreactivity for an apparent high molecular weight phospho-ERK1-containing complex increased in response to synaptic stimulation, but decreased in response to high potassium; p-ERK immunoreactivity at 44/42 kDa increased in both cases. Evidence supporting the conclusion that the band of interest contained ERK1 in a complex, as opposed to it being an unrelated protein crossreacting with antibodies against p-ERK, is that ERK1 (p44 MAPK) and 14-3-3 were electro-eluted from the 160 kDa band cut from a gel. We also found the nuclear complexes to be exceptionally durable, suggesting a role for the crosslinking enzyme, transglutaminase, in its stabilization. In addition, we found other components of the ERK pathway, including MEK, ERK2, p90RSK, and Elk-1, migrating at higher-than-expected weights in brain nuclei. These results describe a novel stable complex of ERK1 in neuronal nuclei that responds differentially to synaptic and depolarizing stimulation, and thus may be capable of mediating gene transcription in a way distinct from the monomeric protein.

B-Raf Associates with and Activates the NHE1 Isoform of the Na+/H+ Exchanger

“…The nature of the complex, whether direct or indirect, is not known. NHE1 is a target for multiple protein kinases, including MAPK (9), and it has been suggested that they may exist as complexes with other members of their pathway (38,39). It is possible that B-Raf is one member of a complex, although this still has to be demonstrated.…”

“…In these experiments we used other His-tagged proteins to confirm that the binding was to NHE1 and not to the His tag on the protein. It was not clear Rsk , Raf-1, PAK5, and 14-3-3 exist as complexes in some cell types (38,39,(47)(48)(49)(50). These complexes can regulate activity of some proteins acting through scaffold proteins (39,50,51).…”

Pathophysiological Consequences of Neuronal α-Synuclein Overexpression: Impacts on Ion Homeostasis, Stress Signaling, Mitochondrial Integrity, and Electrical Activity

“…With the latter antibody, we detected a signal for phospho-Erk1/2 not at the predicted molecular weight of 42/44 kDa, but predominately at a larger size of apparent MW of 160 kDa. A signaling complex of this size containing Erk1/2 has been described in neurons earlier, termed PERC-160 (Lundquist and Dudek, 2006 ).…”

Transglutaminase Inhibition Protects against Oxidative Stress-Induced Neuronal Death Downstream of Pathological ERK Activation

“…Prior elegant work from DeFranco and colleagues revealed a delayed and sustained activation of ERK1/2 owing to redox mediated MKP-1 or PP2A inhibition (Ho et al, 2007). Intriguingly, prior data also suggests that ERK can activate TGs (Akimov and Belkin, 2003) and the transamidating activity may induce heterodimerization of ERK proteins into a complex in the nucleus (Lundquist and Dudek, 2006). We thus examined whether TG activity is downstream of pathological ERK activation in glutathione depletion-induced oxidative stress.…”